The Na,K-ATPase transporters comprise a group of relatively ubiquitous plasma membrane isozymes that use the energy from the hydrolysis of ATP to exchange cytoplasmic Na+ for extracellular K+. Isozyme diversity for the Na,K-ATPase results from the association of different molecular forms, or isoforms of two polypeptides, the catalytic [unreadable] and glycosylated [unreadable] subunits. Among these isoforms, the [unreadable]4 polypeptide has the most restricted pattern of expression, and is selectively present in male germ cells. The [unreadable]4 isoform exhibits several biochemical and functional characteristics that are highly unique and essential to sperm function. Importantly, [unreadable]4 is necessary for normal motility of spermatozoa. Supporting its biological importance, we have shown that [unreadable]4 undergoes significant developmental changes, and that its level of expression and activity increase dramatically after sperm meiosis. Once released, spermatozoa continue to undergo critical maturational changes that are essential for the cells to acquire their fertilizing ability. During this process, called capacitation, we found [unreadable]4 activity to be up-regulated. While these results suggest an important role for [unreadable]4 in sperm function, the precise mechanism(s) by which this protein contributes to sperm capacitation to regulate sperm fertility has yet to be fully defined. Our overall goal in this research is to determine the mechanism(s) by which the activity and ion transport function of the [unreadable]4 isoform is regulated and the relevance of this event in sperm capacitation. Our central hypothesis is that the [unreadable]4 isoform plays an important role in sperm physiology to maintain the ion gradients that control pH, membrane potential and motility changes that are hallmarks of sperm capacitation. Experiments are designed to identify the factors and mechanisms responsible for up-regulation of [unreadable]4 activity, and to determine the contribution of these events to sperm capacitation. The results will have an important positive impact, because they will help to define the molecular basis of sperm capacitation and will contribute to future pharmacologic approaches that will target the [unreadable]4 isoform to control male fertility.